DESCRIPTION (adapted from the applicant's description): Cardiac gap junctions are vital to conduction of the cardiac action potential since they form a low resistance intercellular pathway for ionic current flow between cardiac cells. The mechanism(s) for ion permeation and the connexin protein domain that forms the hydrophilic pore remains unknown. The main goal of this research project is to determine how cations and anions permeate through different connexin channel pores and identify the protein domains that form the ionic permeation pathway. The differential ionic permeability of connexin-specific channels will be determined by varying the concentration and cation/anion composition of permeant ionic salts. Relative ionic permeability ratios will be determined from the diffusion potential developed under asymmetric (biionic) salt conditions and the concentration-dependence or counterion-dependence of these values will distinguish between three different hypotheses for cation and anion permeation (electrostatic forces, countercurrent flux, cotransport of ions) through a pore. The mechanism of ionic blockade by tetraalkylammonium (TAA) and polyamines within the connexin pore will be determined as a measure of the number of sites required and the fraction of the voltage field sensed by the blocking ions. In addition, the conductance and ionic permeability properties of heterotypic connexin channels will be determined. Chimeric and site-directed mutant connexins from two connexins of different conductances and ionic permeabilities will be studied to determine which molecular domain(s) form the channel pore.